透明质酸水凝胶三维培养促进人胚胎干细胞向红系细胞诱导分化  

Three dimensional culture in hyaluronic acid based hydrogel promotes proliferation of erythroid cells derived from hESCs

作　　者：曾宪波[1] 毛斌[1] 赖默温 潘旭[1] 孙文翠[1] 周琼秀[1] 马峰[1] ZENG Xianbo MAO Bin LAI Mowen PAN Xu SUN Wencui ZHOU Qiongxiu MA Feng.(Institute of Blood Transfusion, Chinese Academy of Medical Sciences ＆ Peking Union Medical College)

机构地区：[1]中国医学科学院北京协和医学院输血研究所,四川成都610052

出　　处：《中国输血杂志》2017年第1期12-16,共5页Chinese Journal of Blood Transfusion

基　　金：国家自然科学基金(面上项目5001042804)

摘　　要：目的探讨采用透明质酸材料的三维培养方法将人胚胎干细胞(h ESCs)来源造血干/祖细胞向红系细胞诱导分化的效果。方法将hESCs与小鼠主动脉-性腺-中肾基质细胞(AGM-S3)共培养(37℃、5%CO_2)12 d的细胞(1.0×10~6)分别种植进海藻酸钠水凝胶、胶原蛋白水凝胶、透明质酸水凝胶内,并以悬浮法培养h ESCs作对照。以流式细胞仪检测细胞表面抗原表达,比较不同材料建立的三维培养的优劣,优化红系诱导分化的三维培养体系。结果海藻酸钠组内细胞培养10 d的GPA^+细胞为2.3%,GPA^+CD36+细胞仅为0.9%而悬浮培养分别为15.8%和6.1%。胶原蛋白组内细胞培养10 d的GPA^+细胞为11.1%,GPA^+CD36+细胞为5.2%,而悬浮培养分别为35.7%和27.7%。透明质酸组内红系细胞GPA^+细胞量和比率分别为2.06×105个和19.6%,而悬浮培养的红系细胞GPA^+细胞量和比率分别为1.62×10~5个和15.7%,透明质酸组是悬浮培养的1.27倍,其中悬浮培养GPA^+CD36-为9.7%,而透明质酸水凝胶为GPA^+CD36-为13.1%。结论与悬浮培养相比,海藻酸钠水凝胶和胶原蛋白水凝胶未表现出明显的扩增红系细胞的效果。透明质酸凝胶建立的三维培养方法利于红系细胞生长,该培养方法诱导分化hESCs得到的红系细胞成熟度提高。Objective To establish a three-dimensional culture system in order to culture erythroid cells derived from hematopoietic stem / progenitor cells（ HSPCs） generated from hESCs / AGM-S3 co-cultured in vitro. Methods HSPCs（1. 0× 10^6） from hESCs/AGM-S3 co-cultured（37℃,5% CO2） for 12 days were seeded into alginate hydrogels,collagen hydrogels and hyaluronic acid（ HA） hydrogels respectively,and all hydrogels were compared with suspension culture of hESCs. The proliferation of erythroid cells in different materials were then compared by flow cytometry in order to select the most suitable materials for three-dimensional culture of erythroid cells. Results GPA^＋cells were 2. 3% and GPA^＋CD36^＋cells were 0. 9% in alginate hydrogels,however those cells were 15. 8% and 6. 1%,respectively,in suspension culture. In the collagen group,GPA^＋cells were 11. 1% and GPA^＋CD36^＋cells were 5. 2%,but cells in suspension culture were35. 7% and 27. 7%,respectively. The amount and ratio of GPA^＋cells was 1. 62 × 105 and 15. 7%,respectively,in suspension culture,while it was 2. 06 × 10^5 and 19. 6% in HA. The GPA^＋erythroid cells produced in the hyaluronic acid hydrogel was 1. 27 times higher than in the suspension culture. Among GPA^＋cells,GPA^＋CD36^-cells were 9. 7% in the suspension culture,while HA hydrogels reached 13. 1%. Conclusion Alginate hydrogels and collagen hydrogel are not suitable for the growth of erythroid cells,but hyaluronic acid hydrogel favors erythroid cell maturation.

关 键 词：人胚胎干细胞 红系祖细胞 透明质酸 三维培养 造血干/祖细胞 诱导 分化 

分 类 号：R457.7[医药卫生—治疗学] Q813.11[医药卫生—临床医学]